RESEARCH ARTICLE Exploitation of new approach to control of environmental pathogenic bacteria causing bovine clinical mastitis using novel anti-biofilm nanocomposite Asmaa N. Mohammed 1 & Abeer M. Radi 2 & Rehab Khaled 3 & Fatma I. Abo El-Ela 2 & Amna A. Kotp 3 Received: 20 February 2020 /Accepted: 7 July 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020 Abstract New approaches are required for prevention and control of biofilm-producing bacteria and consequently mitigating the health problems of bovine clinical mastitis. This work designed to determine prevalence rates of biofilm-producing bacteria that causing bovine clinical mastitis and evaluate the anti-biofilm effectiveness of novel nanocomposite of zinc–aluminum layered double hydroxide intercalated with gallic acid (GA) as chelating agent (Zn-Al LDH/GA) on the prevention and control of environmental pathogenic bacteria; Escherichia coli (E. coli), Klebsiella pneumoniae (K. pneumoniae), Staphylococcus aureus (S. aureus), and Coagulase-negative staphylococci (CNS), besides Listeria monocytogenes (L. monocytogenes) and assess the ability to use as an antimicrobial agent, and/or sanitizer for milking equipment. All samples (n = 230) involved clinical mastitis cow’s milk (n = 50) beside environmental samples (n = 180) were collected then examined for isolation and identification of bacterial pathogens. Zn- Al LDH/GA nanocomposite was synthesized using co-precipitation method, then characterized by Fourier-transform infrared spectroscopy (FT-IR); X-ray diffraction (XRD); field emission scanning electron microscopy (FESEM); high-resolution trans- mission electron microscopy (HRTEM); thermogravimetric analysis (TGA); differential thermal analysis (DTA); zeta potential; DLS analysis; and Brunauer, Emmett, and Teller (BET) surface area. The anti-biofilm activity of nanocomposite against mastitis- causing bacteria was detected using the broth micro-dilution and disc-diffusion assay. Results, the minimum concentration of Zn- Al LDH/GA that inhibited the growth of gram-positive and negative bacteria, were 312–625 and 5000 μg/mL, respectively. The LD 50 of Zn-Al LDH/GA was determined in mice at 1983.3 mg/kg b.wt. As a conclusion, Zn-Al LDH/GA nanocomposite proved its efficiency as an antimicrobial agent and/or sanitizer used for cleaning of milking equipment, due to it could inhibit the growth and multiplication of potentially pathogenic bacteria that causing clinical mastitis and its formation of biofilm on the milking equipment. Zn-Al LDH/GA was found to use under varying pH conditions compared with other commercial sanitizer used besides the formation of nanocomposite increases the material stability. Keywords Biofilm-forming bacteria . Environmental contamination . Bovine mastitis . Zinc aluminum–gallic acid . Surface study . Antimicrobial effectiveness . LD 50 Introduction Bovine mastitis is an intramammary disease that causes inflammation of the mammary gland of dairy cows (Bradley 2002; Thompson-Crispi et al. 2014) and it is considered one of the main economic problems facing the global dairy industry and threaten animal welfare, milk quality, and public health (Zadoks and Fitzpatrick 2009; De Vliegher et al. 2012). Biofilm-forming bacteria are the principle cause of intramammary clinical mastitis in lactating cows which involves Staphylococcus aureus, Escherichia coli, and Streptococcus uberis (De Vliegher et al. 2012). Responsible Editor: Philippe Garrigues Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11356-020-10054-1) contains supplementary material, which is available to authorized users. * Asmaa N. Mohammed asmaa.mohamed2@vet.bsu.edu.eg 1 Department of Hygiene, Zoonoses and Epidemiology, Faculty of Veterinary Medicine, Beni-Suef University, Beni-Suef 62511, Egypt 2 Department of Pharmacology, Faculty of Veterinary Medicine, Beni-Suef University, Beni-Suef 62511, Egypt 3 Department of Chemistry, Faculty of Science, Beni-Suef University, Beni-Suef, Egypt Environmental Science and Pollution Research https://doi.org/10.1007/s11356-020-10054-1